KR100353143B1 - Plating medium for the isolation and detection of E. coli O157:H7 - Google Patents

Abstract

The present invention relates to a solid plate medium for the detection and separation of E. coli O157: H7 in a sample, specifically, the medium for detecting E. coli O157: H7 of the present invention comprises: (1) a component for growing microorganisms; (2) agar; (3) sorbitol; (4) chromogenic substrate for beta-galactosidase; (5) chromogenic substrates for beta-glucuronidase; (6) glucuronide components that promote beta-glucuronidase color development reactions; And (7) an ingredient containing erythritol and the like, which inhibits the growth of bacteria other than Escherichia coli O157: H7; pH indicators; And a carbon source not used by E. coli O157: H7. The medium for detecting E. coli O157: H7 of the present invention is not only excellent in selectivity for E. coli O157: H7, but also economical by using an expensive coloring substrate in a minimum amount.

Description

Plating medium for the isolation and detection of E. coli O157: H7}

The present invention relates to a flat medium composition for the detection and separation of Escherichia coli O157: H7. Specifically, the present invention (1) a component for the growth of microorganisms; (2) agar; (3) sorbitol 5-20 g / L; (4) chromogenic substrate for beta-galactosidase; (5) chromogenic substrates for beta-glucuronidase; (6) glucuronide components that promote beta-glucuronidase color development reactions; And (7) E. coli O157: H7 detection and separation medium composition containing 5-20 g / L of erythritol.

E. coli O157: H7 has been identified as a globally important food poisoning bacterium, with the first known cause of the onset of hemorrhagic colitis in 1982, with thousands of infected in Japan and more than 20,000 infected and more than 250 deaths annually in the United States. lost. E. coli O157: H7 is Campylobacter (Campylobacter) bacteria, Salmonella (Salmonella) bacteria, Shigella (Shigella) and bacteria which is a major source of hemorrhagic colitis, along with bacteria, from about 5% of hemorrhagic colitis caused by the fungus is hemolytic-uremic As the disease progresses to a haemolytic uremic syndrom (HUS), various complications and severe deaths have emerged. Therefore, the importance of an effective diagnostic method for preventing or treating infection of E. coli O157: H7 has emerged.

Escherichia coli O157: H7 is known to spread mainly through meat, dairy products, and drinking water. Therefore, there is a need for a method for quickly and economically confirming the presence of E. coli O157: H7 in samples such as food. Conventional methods for testing large samples for E. coli O157: H7 include culturing the sample with a combination of selective liquid or solid medium to isolate the bacterium suspected of E. coli O157: H7, and then confirm this by biochemical or immunological methods. will be. For example, in the E. coli O157: H7 detection test published by the Korea Food and Drug Administration, 25g of the sample was homogenized in 225ml mEC medium and incubated for 24 hours at 35 ° C. Plated on 'SMAC medium' and incubated at 35 ° C for 18 hours. Here, the colorless colonies were taken and plated on EMB agar medium and incubated for 24 hours at 37 ° C., and the green metallic glowing colonies were transferred to Brain Heart Infusion Agar and incubated at 35 ° C. for 24 hours. After confirming the identity of E. coli by biochemical test using API 20E or microbial identifier, the serotype is confirmed by using O157 antiserum and H7 antiserum. This process of completely separating and identifying Escherichia coli O157: H7 takes at least 4 to 5 days. To reduce the additional cost and labor associated with taking false positive bacteria in plate cultures, The need to develop more selective media has been suggested.

The most widely used solid plate medium for the separation of E. coli O157: H7 is SMAC medium, which replaces the original carbon source lactose with 1% sorbitol in MacConkey medium (March and Ratnam, J. Clin). Microbiol. , 23: 869-872, 1986). This medium takes advantage of the fact that most E. coli O157: H7 does not use sorbitol as a carbon source.Bacterium using sorbitol generates an acid as a metabolite, and the colonies develop pink due to the action of the pH indicator contained in the medium. Escherichia coli O157: H7, on the other hand, forms a colorless colony. In addition, the bail salt contained in the medium inhibits the growth of the majority of Gram-positive bacteria, thereby conferring gram-negative selectivity against enteric bacteria. However, the strain does not use sorbitol as a carbon source is E. coli O157: H7 in addition to a number of E. coli, Proteus in (Proteus), Aero Pseudomonas genus (Aeromonas), flash Omo eggplant in (Plesiomonas), do not know the Nella in (Morganella), Providencia SMAC medium has a disadvantage in that the number of false positive bacteria is high because there are so many microorganisms such as Providencia .

There are several studies on improved media based on SMAC media. CT-SMAC medium is a medium containing cepicime and tellurite in SMAC medium. Sepicsim inhibits the growth of proteus and tellulite is Escherichia coli. Inhibits the growth of E. coli and sorbitol-negative bacteria other than O157: H7 (Zadik et al.,J. Med. Microbiol., 39: 155-158, 1993). On the other hand, CR-SMAC medium contains the sepxime and rhamnose in the SMAC medium.E. Coli O157: H7 does not use lamnose on plate media, while about 60% of sorbitol-negative E. coli is lamnose. Acid can be generated as a carbon source, reducing the number of false-positive bacteria (Chapman et al.,J. Med. Microbiol., 35: 107-110, 1991). In addition, it is known that the medium added with novobiocin suppresses the growth of various kinds of bacteria other than Escherichia coli O157: H7 to enhance the selectivity of the SMAC medium. These improved media are more selective than SMAC media but still have a high number of false positive bacteria.

On the other hand, the selectivity can be enhanced by including a color substrate for a specific enzyme in the medium. More than 97% of Escherichia coli produce beta-glucuronidase, whereas E. coli O157: H7 does not produce beta-glucuronidase, so 4-methylumberrirylglucu in medium Ronide (4-methylumbelliferyl glucuronide, MUG) or 5-bromo-4-chloro-3-indolyl-beta-di-glucuronide (5-bromo-4-chloro-3-indolyl-β-D- Inclusion of chromogenic substrates such as glucuronide can reduce the number of false-positive bacteria (Thompson et al. , J. Clin. Microbiol. , 28: 2165-2168, 1990; Okrend et al. , J. Food Protect. , 53: 941-). 943, 1990). This is particularly useful for eradicating Escherichia coli, which does not utilize sorbitol other than Escherichia coli O157: H7.

Unlike non-coli bacteria, colliform bacteria are characterized by metabolism of lactose by producing beta-galactosidase, which is characterized by 5-bromo-4-chloro- in the medium for detecting E. coli O157: H7. 3-indolyl-beta-di-galactopyranoside (5-bromo-4-chloro-3-indolyl-β-D-galactopyranoside) or doxyl-beta-di-galactopyranoside (indoxyl-β Inclusion of chromogenic substrates for beta-galactosidase such as -D-galactopyranoside) can improve the selectivity of media by distinguishing non-coliform bacteria whose biochemical properties are similar to E. coli O157: H7 (JN Roth et. al , USP 5,726,031; B. Bochner, WO 96/40861; L. Restaino, WO 98/1152).

Currently commercial media developed to identify and isolate E. coli O157: H7 contain the chromogenic substrate. Merco's Fluorocult E. coli 0157: H7 medium is a commercial medium containing sorbitol as the carbon source and MUG. Biolog's Rainbow O157 medium contains a substrate for beta-galactosidase and a substrate for beta-glucuronidase, while Chromagar's Chromaga O157 medium is alpha-galactosidase. And chromogenic substrates for glucosidase, and glucuronidase. However, most of these commercial media have a disadvantage of being very expensive due to the large amount of expensive color substrates.

Accordingly, the present inventors have completed the present invention by developing a medium composition excellent in economic efficiency by minimizing the use of an expensive color substrate, which is excellent in detecting ability against E. coli O157: H7, but increases the price of the existing E. coli O157: H7 detection medium.

It is an object of the present invention to provide a composition of a medium for detecting Escherichia coli O157: H7. In particular, the present invention aims to provide a medium composition having excellent selectivity against E. coli O157: H7 while minimizing the use of color substrates in order to improve the economical disadvantages of existing media.

Hereinafter, the present invention will be described in detail.

The medium of the present invention is a solid flat medium for identifying and separating E. coli O157: H7 in a sample. The medium is inoculated with the medium of the present invention and incubated for a predetermined time, and the sample is observed by observing the color development of the colony formed on the medium. To detect E. coli O157: H7.

The basic components of the medium of the present invention are (1) components for the growth of microorganisms; (2) agar; (3) sorbitol 5-20 g / L; (4) chromogenic substrate for beta-galactosidase; (5) chromogenic substrates for beta-glucuronidase; (6) glucuronide components that promote beta-glucuronidase color development reactions; And (7) erythritol 5-20 g / L and the like. In particular, (6) and (7) of the above components are important features of the medium provided by the present invention, serves to enhance the color development function in the medium and is a new component that has not been used in the existing E. coli O157: H7 detection medium. In addition, unlike conventional media, when using a mixture of two or more coloring substrates (4) for beta-galactosidase can further increase the selectivity.

In addition, the medium composition of the present invention, in addition to the above basic components, ingredients that inhibit the growth of bacteria other than Escherichia coli O157: H7, pH indicators, E. coli O157: H7 can not be used by the other sources of potential carbon source By further including the same, the selectivity of the medium can be further increased.

Hereinafter, each component will be described in detail.

In the present invention as a solid plate medium component for the growth of microorganisms, peptone 5 ~ 15g / L as a nitrogen source, sodium chloride 1 ~ 5g / L for the osmotic pressure and minerals provided, yeast extract 0.1 ~ as a source of various vitamins, amino acids, trace elements 2 g / L, and agar 10-20 g / L is preferable as the solidifying component of the medium. In addition, as is well known in the art of microbial culture, components which may replace or additionally use these components may be used.

Coliform bacteria, including E. coli, produce beta-galactosidase, and thus, many non-coliform bacteria can be distinguished and eliminated by including a color substrate for beta-galactosidase in the medium. The medium of the present invention is a 5-bromo-3-indolyl-beta-di-galactopyranoside (5-bromo-3-indolyl-β-D-galactopyranoside), 5- as a beta-galactosidase substrate. Bromo-4-chloro-3-indolyl-beta-di-galactopyranoside (5-bromo-4-chloro-3-indolyl-β-D-galactopyranoside), dodecyl-beta-di-galacto Group consisting of pyranoside (indoxyl-β-D-galactopyranoside) and 4-chloro-3-indolyl-beta-di-galactopyranoside (4-chloro-3-indolyl-β-D-galactopyranoside) It contains the component selected from.

As a carbon source that E. coli O157: H7 cannot use, the medium of the present invention contains 5 to 20 g / L of sorbitol. In the bacterium that can use sorbitol as a carbon source among the colloidal bacteria, metabolism of sorbitol inhibits the beta-galactosidase color development due to acid generation, inhibition of catabolism due to selective metabolism of carbon sources, or other mechanisms. In E. coli O157: H7, such inhibition of beta-galactosidase color development does not occur, so it is possible to distinguish between E. coli O157: H7 and other colloidal bacteria according to the intensity of color development.

Unlike the conventional medium, the medium of the present invention contains 5 to 20 g / L of erythritol as a carbon source. The present inventors confirmed that the addition of erythritol in the medium significantly enhanced the color development of E. coli O157: H7 by the beta-galactosidase substrate, while the color concentration of other colloidal bacteria was relatively weak. Addition of erythritol to the medium is an important feature of the medium of the present invention, it can be used to increase the color concentration without increasing the concentration of the color substrate. Therefore, the use of the medium of the present invention containing erythritol may increase the difference in color development concentrations of Escherichia coli O157: H7 and other colloidal bacteria, thereby significantly improving the selectivity to E. coli O157: H7.

We also include two or more chromogenic substrates for beta-galactosidase with the use of sorbitol and erythritol, unlike the prior art, which uses one chromogenic substrate for beta-galactosidase. In the case of E. coli O157: H7 and other collie-type bacteria, the difference in the color development can be further increased. Therefore, the use of the beta-galactosidase color development substrate in the present invention is not limited to one type, two or more types of substrates selected from the above color development substrates may be used in combination of a ratio of 1: 1 to 1: 5. Here, the ratio of 1: 1 to 1: 5 means the ratio between the component having the minimum amount and the component having the maximum amount among the chromophore substrates. In one embodiment of the present invention, 5-bromo-3-indolyl-beta-di-galactopyranoside and 4-chloro-3-indolyl-beta-di-galacto as beta-galactosidase substrates. Pyranosides each contain 50 mg / L, which shows a color development of E. coli O157: H7 when compared to medium containing 100 mg / L of 5-bromo-3-indolyl-beta-di-galactopyranoside. Similarly, the development of other colloidal bacteria is relatively weak. Thus, despite the use of the same total concentration of chromogenic substrates, a clear improvement in selectivity for E. coli O157: H7 is possible.

To enhance the differentiation of Escherichia coli O157: H7 from other Escherichia coli, the medium also includes a chromogenic substrate for beta-glucuronidase. In the present invention, 4-methylumbelliferyl glucuronide (MUG) 20-200 mg / L, 5-bromo-4-chloro-3- as a coloring substrate for beta-glucuronidase. Indolyl-beta-di-glucuronide (5-bromo-4-chloro-3-indolyl-β-D-glucuronide) 20-200 mg / L, 5-bromo-6-chloro-3-indolyl- Beta-di-glucuronide (5-bromo-6-chloro-3-indolyl-β-D-glucuronide) 20-200 mg / L, or 6-chloro-3-indolyl-beta-di-glucuro If you prefer to use 20 ~ 200mg / L of amide (6-chloro-3-indolyl-β-D-glucuronide), but in addition, if it is a substrate that causes color development that is different from the color reaction of beta-galactosidase, Can be used.

The medium also contains methyl-beta-di-glucuronide, phenyl-beta-di-glucuronide, or 4-nitrophenyl-beta in addition to the chromogenic substrate for beta-glucuronidase described above. -20-200 mg / L of glucuronide, such as di-glucuronide. This is another feature of the present invention, which is distinguished from the conventional E. coli O157: H7 detection medium, and when glucuronide is included, the reaction of the beta-glucuronidase described above is promoted, thereby greatly improving the color intensity. Therefore, a similar level of color development can be obtained by reducing the concentration of color substrate by about half. Since the cost of the beta-glucuronidase chromogenic substrate in the conventional medium for detecting E. coli O157: H7 containing the beta-glucuronidase chromogenic substrate is more than 50% of the total raw material price of the medium, Including such low-cost glucuronide in the medium is an important improvement that can increase the economics of the medium.

On the other hand, in order to increase the selectivity of the medium of the present invention may include a component that selectively inhibits the growth of bacteria other than E. coli O157: H7. Preferred ingredients in the present invention are 0.5 to 3 g / L of a bile salt for inhibiting the growth of Gram-positive bacteria and 5 to 100 mg / L of novobiocin for the growth inhibition of sorbitol-negative bacteria other than Escherichia coli O157: H7. And tellurite (Potassium tellurite) 0.1 ~ 5mg / L, Sepiksim 0.01 ~ 0.5mg / L and the like, in addition to the above components can be included in the medium to inhibit the growth of bacteria other than E. coli O157: H7.

A pH indicator may be included in the medium of the present invention to enhance the distinction between E. coli O157: H7 that does not use sorbitol and strains that use sorbitol. Phenol Red 5-100 mg / L is preferable as a pH indicator, but in addition, color distinguished from the beta-galactosidase color development reaction under acidic conditions such as bromothymol blue and neutral red. If the pH indicator to form a can be used in the medium of the present invention.

E. coli O157: H7 is not available as a carbon source other than sorbitol, but additional inclusions in the medium that may be potentially available by other bacteria may increase selectivity to E. coli O157: H7. Specifically, cellobioses may be used, and in addition, any carbon component not used by E. coli O157: H7 may be used. In addition to cellobiotics, there are other low-speed carbon components that are not used by Escherichia coli O157: H7 in flat media, and include, in addition to cellobiose, lamnose, salicylic acid, esculin, adonitol, and inositol (S. Ratnam et al , J. Clin. Microbiol., 26: 2006-2012, 1988).

Hereinafter, the present invention will be described in detail by way of examples.

However, the following examples are merely to illustrate the invention, but the content of the present invention is not limited to the following examples.

Example 1 Erythritol Containing Effect

To investigate the effect of erythritol contained in the medium on the color reaction of beta-galactosidase, medium # 11, # 12, # 13 and # 14 were prepared according to the compositions shown in Table 1 . For the production of the medium, the components other than the enzyme substrate were dissolved in distilled water and sterilized at 121 ° C. for 20 minutes. The enzyme substrate was dissolved in an appropriate amount of dimethylformamide, and the sterilized medium was cooled to 60 ° C. or lower, and then added. The medium thus prepared was poured into a petri dish to prepare a solid plate medium.

Badge # 11 Badge # 12 Badge # 13 Badge # 14 Common ingredient Bacterium peptone 15g / L, sorbitol 10g / L, bail salt 1.5g / L, sodium chloride 5g / L, agar 15g / L 5-Bromo-4-chloro-3-indolyl-beta-di-galacpyranoside (mg / L) 100 50 100 50 Erythritol (g / L) - - 10 10

Strains used in this Example include E. coli 0157: H7 (ATCC 43890 or 43894); E. coli O157: H19 ( E. coli Reference Center culture strain, Pennsylvania, USA, refererence name: A2), E. coli O26: H46 ( E. coli Reference Center culture strain, Pennsylvania, USA, refererence name: 5306-56), E. coli # 002 (not clinically isolated from O157, clinically isolated from Yonsei Severance Hospital), Proteus mirabilis (Yeonsei Severance Hospital), Salmonella typhimurium , Yersinia enterocoli Ticas ( Yersinia enterocolitica ).

The liquid culture medium in which the strain was cultured was diluted to an appropriate concentration with saline, smeared on a solid plate medium, and the colonies formed after 20 hours of incubation at 37 ° C were observed. The colony color of the strains tested in each medium is shown in Table 2 .

As can be seen from the results of Table 2 , it can be seen that the beta-galactosidase color development was increased when erythritol was included in the medium. In addition, it was difficult to distinguish between colonies in E. coli O157: H7 and other strains due to the weak color development of colonies in medium # 11 and medium # 12.However, in medium # 13 and medium # 14 containing erythritol, the color development was clear. Differentiation between other strains is easier.

Strain Badge # 11 Badge # 12 Badge # 13 Badge # 14 E. coli 0157: H7 (ATCC 43890) Light green Very light green Dark teal Teal E. coli 0157: H7 (ATCC 43894) Light green Very light green Dark teal Teal E. coli O157: H19 Very light green White Teal Light green E. coli O26: H46 White White White White E. coli # 002 (clinical isolates, not O157) Light green Very light green Teal Light green Proteus mirabilis Colorless Colorless Colorless Colorless Salmonella typhimurium White White White White Yersinia enterocolitica White White White White

<Example 2> Effect of using a plurality of beta-galactosidase color substrate

In this example, the color development of the colonies according to the use of a plurality of color substrates for beta-galactosidase was investigated. First, media # 21, # 22, # 23, # 24 and # 25 were prepared according to the compositions shown in Table 3 .

Badge # 21 Badge # 22 Badge # 23 Badge # 24 Badge # 25 Common ingredient Bacterium peptone 15g / L, sorbitol 10g / L, erythritol 10g / L, bail salt 1.5g / L, sodium chloride 5g / L, agar 15g / L 5-Bromo-4-chloro-3-indolyl-beta-di-galactopyranoside (mg / L) 100 - - 50 - 5-Bromo-3-indolyl-beta-di-galactopyranoside (mg / L) - 100 - - 50 4-Chloro-3-indolyl-beta-di-galactopyranoside (mg / L) - - 100 50 50

Specific production method of the medium and smearing of the strain was performed in the same manner as in Example 1, Table 4 shows the color of the population of the strain in each medium.

Strain badge Badge # 21 Badge # 22 Badge # 23 Badge # 24 Badge # 25 E. coli 0157: H7 (ATCC 43890) Dark teal Black Blue White Dark teal Black Blue E. coli 0157: H7 (ATCC 43894) Dark teal Black Blue White Dark teal Black Blue E. coli O157: H19 Teal grey White White White E. coli O26: H46 Greenish white grey White White White E. coli # 002 Teal grey White Light green Light gray Proteus mirabilis Colorless Colorless Colorless Colorless Colorless Salmonella typhimurium White White White White White Yersinia enterocolitica White White White White White

As a result of this experiment, color development did not occur in medium # 23 using 4-chloro-3-indolyl-beta-di-galactopyranoside alone. However, the color development concentration of E. coli O157: H7 is 4-chloro-3-indolyl-beta-di-galactopyranoside to 5-bromo-4-chloro-3-indolyl-beta-di-galactopy. 5-bromo-4- for medium # 24 in combination with ranoside 50 mg / L or for medium # 25 in combination with 5-bromo-3-indolyl-beta-di-galactopyranoside 50 mg / L Medium # 21 using chloro-3-indolyl-beta-di-galactopyranoside 100 mg / L or medium using 5-bromo-3-indolyl-beta-di-galactopyranoside 100 mg / L Equal to or slightly weaker than color development at # 22. On the other hand, the intensity of the development of strains other than E. coli O157: H7 in medium # 24 and medium # 25 was generally weaker than that in medium # 21 and medium # 22. It can be seen that the selectivity between: H7 and other strains can be improved.

Example 3 Color Reaction of Beta-glucuronidase with Inclusion of Glucuronide

In this example, the color reaction of beta-glucuronidase according to glucuronide contained in the medium was investigated. First, media # 31, # 32, # 33, and # 34 were prepared according to the compositions shown in Table 5 .

Badge # 31 Badge # 32 Badge # 33 Badge # 34 Common ingredient Bacterium peptone 15g / L, sorbitol 10g / L, bail salt 1.5g / L, sodium chloride 5g / L, agar 15g / L 5-Bromo-4-chloro-3-indolyl-beta-di-glucuronide (mg / L) 100 50 100 50 4-nitrophenyl-beta-di-glucuronide (mg / L) - - 100 100

Specific production method of the medium and smearing of the strain was carried out in the same manner as in Example 1, Table 6 shows the color of the population of the strain in each medium.

Strain Badge # 31 Badge # 32 Badge # 33 Badge # 34 E. coli O157: H7 (ATCC 43890) White White White White E. coli O157: H7 (ATCC 35150) White White White White E. coli O157: H7 (ATCC 43894) White White White White E. coli O157: H19 Teal Light green Dark teal Teal E. coli O26: H46 Teal Light green Dark teal Teal E. coli 002 Teal Light green Dark teal Teal Proteus mirabilis Colorless Colorless Colorless Colorless Salmonella typhimurium White White White White Yersinia enterocolitica White White White White

As can be seen from the results of Table 6 , the color reaction of 5-bromo-4-chloro-3-indolyl-beta-di-glucuronide was obtained from 4-nitrophenyl-beta-di-glucuronide. Significantly enhanced by inclusion, the color development of non-O157: H7 Escherichia coli in medium # 34 containing 50 mg / L of 5-bromo-4-chloro-3-indolyl-beta-di-glucuronide was reduced to 5-bromo. It is similar or stronger in color development in medium # 31 containing 100 mg / L of parent-4-chloro-3-indolyl-beta-di-glucuronide.

Example 4 Composition of E. coli O157: H7 Detection Medium

Table 7 to Table 9 show the composition of the medium that can be effectively used to detect E. coli O157: H7 as a specific example that can be implemented from the configuration of the present invention.

ingredient density Bakto peptone 10 g / L Sorbitol 10 g / L Erythritol 10 g / L Sodium chloride 5 g / L Weil Salt # 3 1.5 g / L Yeast extract 0.5 g / L Agar 15 g / L Phenolic red 25 mg / L Private Room-Beta-D-Galactopyranoside 150 mg / L

ingredient density Bakto peptone 10 g / L Sorbitol 10 g / L Erythritol 10 g / L Sodium chloride 5 g / L Weil Salt # 3 1.5 g / L Yeast extract 0.5 g / L Agar 15 g / L Phenolic red 25 mg / L 5-Bromo-3-indolyl-beta-di-galactopyranoside 100 mg / L 4-Chloro-3-indolyl-beta-di-galactopyranoside 50 mg / L Cellobiose 10 g / L

ingredient density Bakto peptone 10 g / L Sorbitol 10 g / L Erythritol 10 g / L Sodium chloride 5 g / L Weil Salt # 3 1.5 g / L Yeast extract 0.5 g / L Agar 15 g / L Phenolic red 25 mg / L 5-Bromo-3-indolyl-beta-di-galactopyranoside 100 mg / L 4-Chloro-3-indolyl-beta-di-galactopyranoside 50 mg / L 5-Bromo-4-chloro-3-indolyl-beta-di-glucuronide 75 mg / L 4-nitrophenyl-beta-di-glucuronide 75 mg / L

The medium of the composition of Table 9 above was tested with various strains.

The colony color of each strain is shown in Table 10 . As the results of Table 10 demonstrate, it is easy to distinguish between E. coli O157: H7 and other strains in the medium of the present invention, and thus, excellent selectivity to E. coli O157: H7.

Strain Color of colony E. coli O157: H7 8 species ATCC 35150, 43888, 43889, 43890, 43894 3 clinical isolates ( E. coli Reference Center, Pennsylvania, USA, reference name: 3104-88, C999-87, C7-88) Dark purple Escherichia coli O157: H19 ( E. coli Reference Center, Pennsylvania State University, USA, refererence name: A2) green Escherichia coli O26: H46 ( E. coli Reference Center predisposal strain, Pennsylvania State University, USA, reference name: 5306-56) Light green E. coli JM 109 (ATCC 53323) non-O157 5 isolates of E. coli (Yantae Severance Hospital) Yellow, green or cyan Salmonella paratyphi (Veterinary Quarantine Service) Yellow, white or colorless Salmonella paratyphi (Veterinary Quarantine Service) Salmonella typhi (ATCC 33459) Salmonella typhimurium (Anyang) Salmonella gallinarum (Veterinary Quarantine Service) Salmonella enteritidis (ATCC 13076) Salmonella dublin (Veterinary Quarantine Service) Yersinia enterocolitica (ATCC 23715) yellow Proteus mirabilis (ATCC 14273) Colorless Citrobacter freundii (Yantai Severance Hospital) Greenish yellow transparent Klebsiella pneumoniae (ATCC 10031) White or yellow Serratia marcescens (Yantai Severance Hospital) Colorless Providencia stuartii (Yantai Severance Hospital) Colorless Providencia rettgeri (Yantai Severance Hospital) green Proteus vulgaris (Yantai Severance Hospital) Colorless Morganella morganii (Yantai Severance Hospital) Colorless Acinetobacter anitratus (Yantai Severance Hospital) Colorless Pseudomonas aeruginosa (ATCC 9027) Light green transparent

As described and demonstrated in detail above, the composition of the present invention enables the implementation of a medium with excellent selectivity and economical to E. coli O157: H7.

Claims (13)

A solid plate medium for the growth of microorganisms, comprising: (1) 5-20 g / L of sorbitol; (2) 5-bromo-3-indolyl-beta-di-galactopyranoside, 5-bromo-4-chloro-3-indolyl-beta-di-galactopyranoside, doxyl- Coloring Substrate for One or Two or More Beta-Galactosidases Selected from the Group consisting of beta-di-galactopyranoside and 4-chloro-3-indolyl-beta-di-galactopyranoside 20 ˜200 mg / L; (3) 4-methylumbelliferyl glucuronide (MUG), 5-bromo-4-chloro-3-indolyl-beta-di-glucuronide, 5-bromo- Beta-glucuronidase selected from the group consisting of 6-chloro-3-indolyl-beta-di-glucuronide, and 6-chloro-3-indolyl-beta-di-glucuronide. For chromogenic substrate 20 ~ 200mg / L; (4) beta-glucuro selected from the group consisting of methyl-beta-di-glucuronide, phenyl-beta-di-glucuronide and 4-nitrophenyl-beta-di-glucuronide 20-200 mg / L of glucuronide to promote nidase color reaction; And (5) erythritol 5-20 g / L. A medium composition for detecting E. coli O157: H7. delete According to claim 1, wherein the chromogenic substrate for the beta-galactosidase is 5-bromo-3-indolyl-beta-di-galactopyranoside, 5-bromo-4-chloro-3-indole 2 selected from the group consisting of reel-beta-di-galactopyranoside, indoxyl-beta-di-galactopyranoside, and 4-chloro-3-indolyl-beta-di-galactopyranoside A component of the species, the composition ratio of which is 1: 1 to 1: 5. According to claim 1, wherein the coloring substrate for the beta-galactosidase is 5-bromo-3-indolyl-beta-di-galactopyranoside 20 ~ 200mg / L and 4-chloro-3-indole A composition comprising a combination of reel-beta-di-galactopyranoside 20-200 mg / L. delete delete According to claim 1, wherein in addition to the above-mentioned ingredients, one selected from the group consisting of 0.5-3 g / L of bail salt, 0.1-5 mg / L of tellurite, 5-100 mg / L of Novobiocin and 0.01-0.5 mg / L of Sepiksim Or a component that inhibits the growth of bacteria other than E. coli O157: H7. delete The composition according to claim 1, wherein the composition contains 5 to 100 mg / L of pH indicator selected from the group consisting of phenol red, bromocymol blue and neutral red in addition to the component. 10. The composition of claim 9, wherein the pH indicator is 5-100 mg / L phenol red. The method according to claim 1, wherein in addition to the above, cellobio 5-20 g / L, lamnose 5-20 g / L, salicylic 5-20 g / L, esculin 5-20 g / L, adonitol 5-20 g / L, And one or two or more components selected from the group consisting of inositol 5 to 20 g / L. Solid medium for the identification and isolation of Escherichia coli O157: H7, (1) 5-15 g / L peptone, 1-5 g / L sodium chloride, and 0.1-2 g / L yeast extract, as components for the growth of microorganisms (2) 5-bromo-3-indolyl as a chromogenic substrate for sorbitol 5-20 g / L, (3) erythritol 5-20 g / L, (4) beta-galactosidase as a carbon source that E. coli O157: H7 cannot use Beta-di-galactopyranoside 20-200 mg / L and 4-chloro-3-indolyl-beta-di-galactopyranoside 20-200 mg / L, (5) agar 10-20 g / L, (6) 0.5-3 g / L of bail salt, (7) 5-bromo-4-chloro-3-indolyl-beta-di-glucuronide 20- as a chromogenic substrate for beta-glucuronidase. 200 mg / L, (8) methyl-beta-di-glucuronide, phenyl-beta-di-glucuronide, and 4-nitrophenyl-beta-di-glucuronide Composition comprising 20-200 mg / L, (7) 5-100 mg / L phenol red, and (8) 5-20 g / L cellobiose A method for identifying or isolating E. coli O157: H7 in a sample, which is inoculated with one of the medium of claims 1 to 12, incubated for 16 to 48 hours, and the color development of a colony formed on the medium. Method for detecting E. coli O157: H7 in the sample by observing.